Parkinson's disease is caused by selective degeneration and deletion of dopaminergic neurons in mesencephalic substantia nigra. For treating it, the effectiveness of transplanting a fetal mesencephalon tissue that contains a large amount of dopaminergic neurons (or cells having the differentiation potency into dopaminergic neurons), into the brain (striate body) of patients has been verified.
In fact, however, it is impossible to secure plenty of fetal brain tissue for use in ordinary clinics. Therefore, donor cells substitutable for fetal mesencephalon are desired.
For example, it is under investigation to use cells that have been differentiated into dopaminergic neurons from a large number of undifferentiated neural cells, for donor cells for transplantation. In addition, it is also under investigation to use cells having been differentiated from non-neural cells such as ES cells or marrow mesenchymal cells into dopaminergic neurons, for donor cells for transplantation. These cells can be differentiated into the intended dopaminergic neurons, after in vitro expansion, and therefore could be a means for solving the problem of shortage of donors. Moreover, since marrow mesenchymal cells can be safely collected from adults, it is possible to prepare dopaminergic neurons for transplantation from the cells of patients themselves. Accordingly, if those kinds of therapeutic strategy become available, they will solve not only the problem of shortage of donors and the technical problem of rejection against grafts, but also the ethical problem involved in obtaining dopaminergic neurons from aborted babies.
However, the method for the efficient induction of dopaminergic neurons from undifferentiated cell groups is not as yet completely established. In addition, from undifferentiated cell groups, various cells other than dopaminergic neurons are differentiated. Further, there is a risk that undifferentiated cell groups may include cells that will form tumors after transplanted. Accordingly, if dopaminergic neurons that had been differentiated in vitro are intended to be used for transplantation, they must be selectively separated from many kinds of cell groups.
As so mentioned hereinabove, enriched dopaminergic neurons are expected to be useful for graft donor cells for treatment of Parkinson's disease, etc. In addition, the technique of enrichment and/or isolation of dopaminergic neurons is extremely useful for identifying novel proteins and genes that are expressed specifically in these neurons. This is because such proteins and genes are expected to lead to novel drugs for the treatment.
In addition, it is extremely important to identify the factor that induces the in vitro differentiation of dopaminergic neurons from undifferentiated cells. Not only the factor is useful for efficiently inducing dopaminergic neurons from undifferentiated cells, but also the factor itself is expected to lead to novel drugs for the treatment.
As yet, however, no method has been established for isolating dopaminergic neurons from in vivo tissues or from cells being cultured in vitro.
Needless-to-say, not only the method for searching for the factor of in-vitro induction of dopaminergic neurons but also the method necessary for the search, which is for visualizing living dopaminergic neurons, has not been established as yet.
One object of the invention of this application is to provide a method for visualizing living dopaminergic neurons in cells including various types of different cells, to thereby enrich and isolate the dopaminergic neurons to a high purity.
Another object of the invention of this application is to provide the dopaminergic neurons isolated by the method.
Still another object of the invention of this application is to provide a method for identifying a factor that induces the differentiation of dopaminergic neurons from undifferentiated cells.